Methanogenic partner influences cell aggregation and signalling of Syntrophobacterium fumaroxidans

Abstract For several decades, the formation of microbial self-aggregates, known as granules, has been extensively documented in the context of anaerobic digestion. However, current understanding of the underlying microbial-associated mechanisms responsible for this phenomenon remains limited. This study examined morphological and biochemical changes associated with cell aggregation in model co-cultures of the syntrophic propionate oxidizing bacterium Syntrophobacterium fumaroxidans and hydrogenotrophic methanogens, Methanospirillum hungatei or Methanobacterium formicicum. Formerly, we observed that when syntrophs grow for long periods with methanogens, cultures tend to form aggregates visible to the eye. In this study, we maintained syntrophic co-cultures of S. fumaroxidans with either M. hungatei or M. formicicum for a year in a fed-batch growth mode to stimulate aggregation. Millimeter-scale aggregates were observed in both co-cultures within the first 5 months of cultivation. In addition, we detected quorum sensing molecules, specifically N-acyl homoserine lactones, in co-culture supernatants preceding the formation of macro-aggregates (with diameter of more than 20 μm). Comparative transcriptomics revealed higher expression of genes related to signal transduction, polysaccharide secretion and metal transporters in the late-aggregation state co-cultures, compared to the initial ones. This is the first study to report in detail both biochemical and physiological changes associated with the aggregate formation in syntrophic methanogenic co-cultures. Keypoints • Syntrophic co-cultures formed mm-scale aggregates within 5 months of fed-batch cultivation. • N-acyl homoserine lactones were detected during the formation of aggregates. • Aggregated co-cultures exhibited upregulated expression of adhesins- and polysaccharide-associated genes. Graphical abstract Supplementary Information The online version contains supplementary material available at 10.1007/s00253-023-12955-w.


Figure S2 .
Figure S2.Chromatograms of AHLs analyzed in SIM mode on UHPLC-MS/MS.Two concentrations were chosen as an example: the smallest (Z, left) and the highest (T, right).
Figure S6.Principal component analysis of the transcriptome samples from the co-cultures of S. fumaroxidans with M. formicium (A) or M. hungatei (B) at the early-aggregation (orange) or late-aggregation (green) state.

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Figure S12.A normalized differential expression of the S. fumaroxidans genes for the central metabolism in Sf-Mf and Sf-Mh early-(Cycle-1) and late-aggregation (Cycles-20/23) state co-cultures.

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Figure S13.A normalized differential expression of the M. formicicum genes for the central metabolism in Sf-Mf early-(Cycle-1) and late-aggregation (Cycle-20) state co-cultures.Fdh = formate dehydrogenases.

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Figure S14.A normalized differential expression of the M. hungatei genes for the central metabolism in Sf-Mh early-(Cycle-1) and late-aggregation (Cycle-23) state co-cultures.Fdh: formate dehydrogenases, MspA: major sheath protein.

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Figure S15.A normalized differential expression of the S. fumaroxidans genes for the secondary metabolism in Sf-Mf and Sf-Mh early-(Cycle-1) and late-aggregation (Cycles 20/23) state co-cultures.

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Figure S16.A normalized differential expression of the M. formicicum genes for the secondary metabolism in Sf-Mf early-(Cycle-1) and late-aggregation (Cycle-20) state co-cultures.

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Figure S17.A normalized differential expression of the M. hungatei genes for the secondary metabolism (biosynthesis of EPS, metal transporters, regulation of flagella and pili) in Sf-Mh early-(Cycle-1) and lateaggregation (Cycle-23) state co-cultures.

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Figure S18.A normalized differential expression of the M. hungatei genes for the secondary metabolism (chemotaxis and signal transduction) in Sf-Mh early-(Cycle-1) and late-aggregation (Cycle-23) state co-cultures.